Method and apparatus for characterization of clot formation

ABSTRACT

Methods, apparatus and systems for characterizing changes in at least one physical property of soft tissue. A series of acoustic pulses is generated and directed into the soft tissue such that at least one of the pulses is of sufficiently high intensity to induce physical displacement of the tissue. Waves reflected off the tissue, or a flexible member that moves with the tissue, are received and measured to estimate at least one characteristic of the physical displacement induced thereby. Repetition of the generating, receiving and estimating steps provides characterization of the at least one physical property over time. Methods, apparatus and systems for characterizing at least one physical property of blood, by generating a series of acoustic pulses and directing the series of pulses into the blood such that at least one of the pulses is of sufficiently high intensity to induce physical displacement of the blood. Acoustic pulses and/or optical waves reflected from the blood, or a flexible member in contact with the blood that moves with the blood, are received and measured to estimate at least one characteristic of the physical displacement induced thereby.

CROSS-REFERENCE

This application claims the benefit of U.S. Provisional Application No.60/513,264, filed Oct. 22, 2003, which application is incorporatedherein, in its entirety, by reference thereto.

GOVERNMENT RIGHTS

This invention was made with government support under federal grant no.GAAN P200A010433 awarded by the Department of Education. The UnitedStates Government may have certain rights in this invention.

BACKGROUND OF THE INVENTION

Blood coagulation is a delicately regulated process that serves as aprotective mechanism against blood loss due to tissue damage. Overactiveor unregulated coagulation can lead to conditions including myocardialinfarction, stroke, deep vein thrombosis (DVT), and pulmonary embolism.The ability to recognize coagulation disorders and quantify theirseverity is critical for identifying those at risk and implementingappropriate prophylactic treatment. Because of inherent risksaccompanying anticoagulant therapy, such, as hemorrhage or anaphylaxis,it is critical that such therapies be prescribed appropriately (seeAnderson et al., “Best Practices: Preventing Deep Vein Thrombosis andPulmonary Embolism”, Center for Outcomes Research, U. Mass. Med. Ctr,1998, which is hereby incorporated herein, in its entirety, by referencethereto).

Hypercoagulability, or thrombophilia, is an inherited or acquiredcoagulation disorder in which there is either an overactivation ofcoagulation or deficient deactivation of developed thrombus. While anumber of factors within the coagulation cascade such as factor VLeiden, protein C or S deficiency, and antithrombin III deficiency areknown to increase the propensity to clot (see Harris et al. “Evaluationof Recurrent Thrombosis and Hypercoagulability”, American FamilyPhysician, vol. 56 (6), Oct. 15, 1997, which is hereby incorporatedherein, in its entirety, by reference thereto), there is currently adearth of techniques available to quantify these effects clinically. Themethods currently available are mostly biochemical in nature and testfor a specific genetic mutation or abnormal chemical reaction rate, suchas Leiden Factor V. mutation R560Q; Hyperhomocysteinemia MTHFR Mutation;Prothrombin Gene Mutation 20210; Protein C levels; Protein S levels;Activated Protein C activity; antibodies to six phospholipids of theIgM, IgG and IgA classes; Lupus anticoagulant antibody; Russell ViperVenom time; Activated Partial Thromboplastin time; and Prothrombin time;see http://repromed.net/papers/thromb.php which is incorporated herein;in its entirety, by reference thereto. While these tests may providevaluable information, they are unable to determine the coagulation rateof an individual's blood. Furthermore, since the coagulation cascade isexceedingly complex, there are numerous steps in the pathway that mightbe disrupted or inappropriately regulated. However, it is not alwayspossible to determine if these interruptions in the cascade arepredictive of an observable clinical impact on thrombus formation.

Mechanical methods, such as cone and plate viscometry or indentationtesting, provide the most intuitive way to characterize the mechanicalparameters of blood coagulation. However, these approaches are limitedbecause the mechanical forces applied to the forming thrombus candisrupt its delicate structure, and thus disturb the system enough tointerrupt the normal course of coagulation.

Deep vein thrombosis (DVT) refers to the formation of a blood clot in alarge vein of the leg, DVT often results from a lack of movement in theextremities for significant periods of time or from an increasedpropensity to clot due to malignancy, recent surgery or trauma,pregnancy, hormonal agents such as oral contraceptives, or othercontributing causes, see Hirsh et al., “How We Diagnose and Treat DeepVein Thrombosis”, Blood, vol. 99(1), pp. 3102-3110, which is herebyincorporated herein, in its entirety, by reference thereto. If a portionof the thrombus breaks off and travels to the pulmonary vessels, apotentially fatal pulmonary embolism can result. Clinical diagnosiscannot serve as the sole means of DVT diagnosis because many potentiallydangerous venous thrombi are asymptomatic, and many of the symptoms arenot unique to DVT. Current noninvasive methods of diagnosis such asduplex ultrasonography, venography, impedance plethysmography, and MRIcan often detect the presence of a clot, but are limited by an inabilityto determine the stage of development of the clot so identified.Furthermore, these methods must often be used in conjunction withanother diagnostic method or tool such as the d-dimer assay in order tomake a conclusive diagnosis.

Although duplex ultrasonography is favored for the initial investigationof DVT, several groups have also proposed the use of ultrasound toextrapolate parameters related to the formation of DVT. Shung et al, in“Ultrasonic Characterization of Blood During Coagulation”, Journal ofClinical Ultrasound, vol. 12, pp 147-153, 1984 (which is herebyincorporated herein, in its entirety, by reference thereto), have shownthat the increase in echogenicity associated with the formation of athrombus is mostly due to an increase in ultrasonic backscatter. Theyhave also found increases in both the attenuation coefficient and thespeed of sound. Parsons et al. in “Age Determination of ExperimentalVenous Thrombi by Ultrasonic Tissue Characterization”, Journal ofVascular Surgery, vol. 17(3), pp. 470-478, 1993 (which is herebyincorporated herein, in its entirety, by reference thereto), have beenable to differentiate in vivo between clots of varying ages by looking athe slope and intercept of the linear fit of the normalized powerspectrum. Emelianov et al., in “Ultrasound Elasticity Imaging of DeepVein Thrombosis” Proc. IEEE Ultrasonic Symposium, 2000 (which is herebyincorporated herein, in its entirety, by reference thereto), havecharacterized different clinical stages of a thrombus using maps oflocal strain. Their method operates by obtaining baseline radiofrequency (RF) echo data, mechanically compressing the tissue, obtaininga second compressed set of data, and applying signal processing methodsto create maps of local strain. Rubin et al., in “Clinical applicationof sonographic elasticity imaging for aging of deep venous thrombosis:preliminary findings,” Journal of Ultrasound in Medicine, vol. 22, pp.443-8, 2003, which is incorporated herein, in its entirety, by referencethereto, characterizes different clinical stage of a thrombus using mapsof local strain obtained by compressive elastography. Although thetechniques proposed by Parsons et al., Emelianov et al. and Rubin et al.have yielded valuable results, they are primarily focused on ageclassification of DVT and thus are not able to characterize thrombusformation. Furthermore, these techniques do not provide informationabout coagulability and are thus of little or no value in prospectivelyidentifying patients at high risk of forming a blood clot. Furthermore,direct translation of these techniques to benchtop tools is problematicbecause of high variability in measurements taken.

There remains a need for the ability to characterize changes in softtissue, and particularly for characterizing thrombus formation. Thereremain needs for methods, apparatus and systems that can characterizethrombus formation for diagnosis and treatment purposes, and preferablyin a substantially non-invasive manner.

SUMMARY OF THE INVENTION

The present invention provides methods of characterizing at least onephysical property of soft tissue. One such method described hereinincludes generating a series of acoustic pulses and directing them intothe soft tissue to be characterized, wherein at least one of the pulsesis of sufficiently high intensity to induce physical displacement of thetissue. At least one physical property of the tissue is estimated basedon measurement of at least two of the pulses as reflected from the softtissue and/or receiving optical reflections from the soft tissue as thesoft tissue is being physically displaced. The process may be repeatedat least once after passage of a time interval, so that time-based datacan be generated.

An apparatus for identifying changes in at least one physical parameterof a soft tissue over time includes an acoustic wave generator capableof repeatedly generating acoustic pulses of sufficient intensity toinduce measurable physical displacement in the soft tissue; a sensoradapted to sense at least one of optical waves or the acoustic pulsesafter reflection by the soft tissue; a clock governing cycles duringwhich the acoustic pulses are generated and during which sensing of atleast one of the acoustic or optical waves is carried out; and aprocessor that receives input from the sensor and clock and calculatestime-based data characterizing at least one characteristic of thephysical displacement induced.

A method of characterizing at least one physical property of blood isdescribed, including generating a series of acoustic pulses anddirecting the series of pulses into the blood such that at least one ofthe pulses is of sufficiently high intensity to induce physicaldisplacement of the blood; measuring a displacement of the bloodresulting from the induced physical displacement thereof; and estimatingat least one characteristic of the physical displacement based on themeasurement.

Methods of diagnosis of the development stages of clotting aredescribed.

Methods of evaluating effectiveness of anti-clotting treatments aredescribed.

Methods of evaluating effectiveness of pro-clotting treatments are alsodescribed.

These and other advantages and features of the invention will becomeapparent to those persons skilled in the art upon reading the details ofthe methods, apparatus and systems as more fully described below.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A is a schematic representation of the present invention usefulfor in vitro characterization of a soft tissue sample.

FIG. 1B is a modification of the arrangement shown in FIG. 1A in whichan additional device is positioned on a side of the container oppositethe device that is also shown in FIG. 1A.

FIG. 1C schematically illustrates a non-invasive use of the presentinvention.

FIG. 2 is a schematic representation of a system for characterization ofat lest one physical property of soil tissue.

FIG. 3 shows a series of time-displacement curves comparing valuespredicted by a model to values obtained using an embodiment of thepresent, apparatus.

FIG. 4 is a symbolic representation of a modified Voigt model used as amodel to characterize the behavior plotted in FIG. 3.

FIG. 5 is a diagrammatic representation of apparatus for in vitrocharacterization of at least one physical property of soft tissue.

FIGS. 6A, 6B and 6C show a portion of the results obtained fromanalyzing a control solution as described in the Example below.

FIG. 7 shows comparative time constants for convention rheometry uses ascompared to use of the present techniques.

FIG. 8 shows a set of time-displacement curves obtained from one bloodsample along with the accompanying best fit model predictions, asdescribed below in the Example.

DETAILED DESCRIPTION OF THE INVENTION

Before the present methods, apparatus and systems are described, it isto be understood that this invention is not limited to particularembodiments described, as such may, of course, vary. It is also to beunderstood that the terminology used herein is for the purpose ofdescribing particular embodiments only, and is not intended to belimiting, since the scope of the present invention will be limited onlyby the appended claims.

Where a range of values is provided, it is understood that eachintervening value, to the tenth of the unit of the lower limit unlessthe context clearly dictates otherwise, between the upper and lowerlimits of that range is also specifically disclosed. Each smaller rangebetween any stated value or intervening value in a stated range and anyother stated or intervening value in that stated range is encompassedwithin the invention. The upper and lower limits of these smaller rangesmay independently be included or excluded in the range, and each rangewhere either, neither or both limits are included in the smaller rangesis also encompassed within the invention, subject to any specificallyexcluded limit in the stated range. Where the stated range includes oneor both of the limits, ranges excluding either or both of those includedlimits are also included in the invention.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although any methods andmaterials similar or equivalent to those described herein can be used inthe practice or testing of the present invention, the preferred methodsand materials are now described. All publications mentioned herein areincorporated herein by reference to disclose and describe the methodsand/or materials in connection with which the publications are cited.

It must be noted that as used herein and in the appended claims, thesingular forms “a”, “and”, and “the” include plural referents unless thecontext clearly dictates otherwise. Thus, for example, reference to “atransducer” includes a plurality of such transducers and reference to“the curve” includes reference to one or more curves and equivalentsthereof known to those skilled in the art, and so forth.

The publications discussed herein are provided solely for theirdisclosure prior to the filing date of the present application. Nothingherein is to be construed as an admission that the present invention isnot entitled to antedate such publication by virtue of prior invention.Further, the dates of publication provided may be different from theactual publication dates which may need to be independently confirmed.

The present invention provides methods, apparatus and systems forperforming what the present inventors have termed sonorheometry.Sonorheometry provides data about the mechanical properties of softtissue. Furthermore, repeated measurements using sonorheometry enablecharacterization of changing properties over time. Sonorheometry isparticularly well-suited to characterizing blood coagulation. Thepresent invention provides data about the mechanical properties of adeveloping thrombus without disrupting its formation. The methods andtechniques may be non-invasive or carried out in a laboratory settingafter obtaining a sample from a patient, and are based on theapplication of acoustic radiation force to the tissue to becharacterized.

An increased or decreased propensity to clot can be evaluated byobserving the coagulation rate and mechanical characteristics of thedeveloping thrombus at any time during formation. This information mayin turn allow clinicians to assess an individual's clotting behavior andto treat coagulation disorders appropriately. This information may alsobe used to evaluate whether a particular treatment and/or dosage iseffective or needs to be changed, as subsequent testing according to thepresent methods (i.e., after a treatment has been administered) can becarried out to compare the results, thereby indicating the effect of thetreatment.

Referring now to FIG. 1, an assembly 1 is schematically shown that isset up for testing soft tissue according to the present invention. Anacoustic wave generating device 10 is positioned in alignment withcontainer 30 to allow device 10 to irradiate a soft tissue containedwithin container 30. Device 10 may be mounted or fixed at apredetermined distance for the contents of the container 30 to receivefocused acoustical waves from device 10. Thus, device 10 and container30 are oriented to align the emission of acoustic waves from device 10with a sample contained in container 30. Container 30 may be entirelyacoustically transparent, or contains at least one window 32 a that isacoustically transparent and that is aligned with the emission pathwayof device 10. As one non-limiting example; container 30 may include aplastic cuvette having windows 32 a 32 d cut therethrough and coveredwith KAPTON® (polyimide) film or other acoustically transparent film.One knowledgeable in the art will realize that it may be advantageous toplace the acoustic window or windows of the sample container at somenon-perpendicular angle relative to the direction of wave propagation soas to reduce the magnitude of received echoes from the interfaces withthe window(s). Multiple measurements may be performed at the same timeusing an array of sample containers 30, for example. One knowledgeablein the art will recognize that such an array may either consist ofindividual containers, or a single container with multiple samplecompartments. Additionally or alternatively, an array of transducers maybe included in device 10, or an array of devices 10 may be used to makemultiple measurements. Thus, for example, multiple transducers and/ormultiple devices 10 may be provided to analyze multiple samples inparallel, wherein the multiple samples are contained in multipleindividual containers or a single container with multiple samplecompartments.

Assembly 1 may be submerged in a tank of water or other coupling mediumto facilitate transmission of the acoustic waves. Alternatively, device10 (or other acoustic emitter and receiver) may be placed in directcontact with the sample. Still further, device 10 may be adapted todeposit the sample directly in contact therewith, for example placing adrop (or other quantity) of blood on a transducer contained in device 10or other application feature of device 10. In the case where a bath (ofwater or other coupling medium) is provided, the bath may be a constanttemperature bath or other means may be provided to maintain a constantsample temperature. In cases where no bath is used, it may beadvantageous to place the sample in contact with a material ofcontrolled temperature, so as to control the sample temperature. Anotheralternative is the use of device 10 invasively. For example, device 10may be inserted intravascularly and delivered to the location of a stentto characterize any clotting that may be occurring as well ascharacterize the progression or stage of a clot that may be present.Similar intravascular techniques can be applied for identifying and/orcharacterizing clot processes with regard to DVT, as well as for otherclotting events throughout the body, as long as the location isaccessible by catheter or other delivery instrument, for example. Thus,not only are intravascular insertions, deliveries or locations madpossible by the device, but the device may also be positioned at anintracavity location or other location inside of the body.

Device 10 includes an acoustic wave generating source capable ofgenerating one or more pulses, at least one of which is of sufficientintensity to induce measurable physical displacement in the soft tissuecontained in container 30. For example, device 10 may include one ormore piezoelectric transducers capable of generating ultrasonic waves.Alternatively, device 10 may utilize an electric circuit to generaterapid heating and thereby generate acoustic energy. Further alternativesmay be employed for generating acoustic energy, including, but notlimited to: an ultrasonic generator fabricated usingmicroelectromechanical systems (MEMS); a capacitive micromachinedultrasound transducer; a laser used to heat a target material therebygenerating acoustic energy, where the laser may be targeted on apermanent component of the assembly, or on a surface of the sample, forexample. Still further alternatively, a transducer may be incorporatedinto the sample container 30 in lieu of providing it in the device 10,as in a case, for example, where a polymer transducer material such asPVDF may be glued right onto the surface of the sample container 30.

Device 10 further includes at least one sensor capable of measuringdisplacement or deformation induced by the acoustic waves as they areapplied to the soft tissue sample and reflected by the soft tissuesample back to device 10. In this configuration, an ultrasound sensormay be used to track the motion of the sample as induced by at least oneultrasonic wave of sufficient intensity to induce displacement of thetissue. Alternatively, tracking of the motion may be accomplished bymeans other than sensing reflected acoustic waves. For example, opticalcoherence tomography, a focused light interferometer or laser Dopplermay be used to optically sense the displacement of the tissue induced bythe one or more ultrasonic waves. Device 10 may include one or moresensors for carrying out any of these optical methods or such sensorsmay be provided in equipment that is separate from device 10. Likewise,for acoustic sensing, the one or more sensors may be one and the same asthe acoustic wave generator, or may be a separate component(s) and maytake any of the forms described above with regard to the acoustic wavegenerating component. Typically, an ultrasonic transducer may be used toboth apply ultrasonic waves to the soil tissue as well as to senseultrasonic waves reflected back from the tissue. An adjoining processor(not shown in FIG. 1) may be provided to control the timing oftransmission of pulses and of receiving of echoes (reflected pulses) bydevice 10.

FIG. 1B shows an example wherein a second device 10′ is positioned inalignment with device 10, but on the opposite side of container 30compared to the location of device 10. In this example, container 30 maybe entirely acoustically transparent, or contain at least two windows 32a and 32 d that are acoustically transparent and that are aligned withthe emission pathway of device 10 to permit emissions to pass throughboth windows 32 a and 32 d to be received by device 10′. System 1 shownin FIG. 1B, in addition to performing the measurements that the systemof FIG. 1A performs, can also measure acoustic properties, includingspeed of sound and attenuation, which provide indirect measures oftissue microstructure and which may be used for calibration purposes.

According to Torr, “The Acoustic Radiation Force: Am. J. Phys., vol. 52,pp. 402-408, 1984, which is hereby incorporated herein, in its entirety,by reference thereto, acoustic radiation force arises from two sources:“a non zero time-averaged sound pressure in the ultrasonic beam, and themomentum transported by the beam.” Torr argues, and it has been widelyaccepted, that the momentum transfer component of this force dominatesunder most conditions. This momentum transfer results from attenuationof the propagating ultrasound beam via both absorption and scattering.For the case of total absorption the applied radiation force is simply:

F=W/c  (1)

where W is the acoustic power and c is the speed of sound in the medium.In the case of perfect reflection this radiation force is doubled. Inboth cases radiation force acts along the direction of wave propagation.

In biological media absorption and reflection are neither total, norisolated at interfaces. Rather, attenuation and reflection (in the formof scattering) occur throughout volumes of tissue. In these casesradiation force acts as a body force, with the force on a given volumesimply equal to the sum of the force from absorption and that fromscattering. If we assume that scattering in the tissue consists purelyof backscatter, which is of course overly simplistic, then the radiationforce applied to a given volume of tissue is:

F=W _(u) /c+2W _(s) /c  (2)

where W_(u) is the absorbed ultrasound power and W_(s) is the scatteredultrasound power within the volume. If we further simplify byrecognizing that only a fraction of the scattered energy is returned asbackscatter, and that attenuation is dominated by absorption rather thanscattering, then (2) can be simplified as:

F=W _(α) /c=A/cI ₀(e ^(−2αfz) ¹ −e ^(−2αfz) ² )  (3)

where A is the cross sectional area of the volume of interest(perpendicular to the axis of propagation), I₀ is the ultrasoundintensity that would be observed in the absence of attenuation, α is theamplitude attenuation coefficient in Nepers per centimeter per MHz, f isthe ultrasonic center frequency in MHz, and z₁ and z₂ are the ranges ofthe front and back of the volume in units of centimeters.

By utilizing two devices 10 and 10′ (wherein device 10 at least containsan emitter and device 10′ contains at least a sensor for receiving thewaves/pulses that pass through windows 32 a,32 d the system can alsomeasure the waves that pass from device 10 to device 10′ and estimateacoustic properties of the sample being analyzed. Examples of acousticproperties that may be estimated include attenuation, scattering, andspeed of sound during sonorheometry procedures. The data received bydevice 10′ may be used to make predictions/estimations of the appliedradiation force and compare experimentally determined displacements topredicted displacements.

It should be noted that although FIG. 1A shows an example of apparatusfor performing analysis in vitro (such as in a laboratory setting, orfrom a self-operated testing kit, for example) after taking a sample tobe analyzed from a patient and depositing it in container 30,alternatively, the present invention may also be practicednon-invasively, such as by applying acoustic waves from a device 10transdermally through a patient 2 (in vivo) to the targeted tissue to beanalyzed, see FIG. 1C. A single time frame analysis of one or morephysical properties of the tissue may be made, or time series studiesmay be performed by applying the waves transdermally at different timeperiods, using the techniques described herein for the in vitro studies.Of course the in vivo analyses would typically not involveadministration of thrombin or other coagulant to a patient. However timestudies may be done to test the effectiveness of an anti-clottingtreatment regimen for example. Similarly, time studies may be done totest the effectiveness of a pro-clotting regimen given to a patient toincrease the ability of the blood to clot, such as in the case of ahemophiliac, for example. Likewise, the administration of thrombin isnot necessarily required for time studies in vitro, as there are othertechniques that may be substituted to initiate coagulation, such assnake venom, the use of ground glass to initiate coagulation, etc.

Non-invasive applications of the current invention includecharacterizing a stage of development of a blood clot by generating aseries of acoustic pulses and transdermally directing the series ofpulses into the blood such that at ledst one of the pulses are ofsufficiently high intensity to induce physical displacement of theblood, receiving at least two pulses, including at least one pulsereflected from the blood to establish a baseline and another pulsereflected from the blood to estimate at least one characteristic of thephysical displacement induced by the waves. Alternatively, the at leasttwo pulses identified above as being used for establishing baseline andestimating a characteristic resulting from the physical displacement ofthe sample, do not necessarily have to be reflected from theblood/sample. For example, if the sample is contained within membranesthat move with the movement of the blood/sample or in a container 30that is sufficiently flexible (such as a membranous container, forexample) to move with the movements of the blood/sample, then the atleast two pulses could alternatively be those reflected from thesurfaces of the flexible sample container or other membranes placedwithin the sample, as the movement of the sample (e.g., development ofthe clot) will alter the position of the surfaces or membranes.

The at least one estimate may be compared to previously generated datato gauge the stage of development of the blood clot being analyzed. Thepreviously generated data may be reference data, such as generatedacross a larger number of patients and then averaged to determine normalcharacteristics, as well as to find average levels for characterizingdifferent stages of clotting for example. Optionally, one or morealgorithms, techniques or statistical processes may be applied to the atleast one estimate to correct for attenuation, scatter and/or othervariables before making comparisons to the previously generated dataand/or database. Additionally, or alternatively, the prior data orpreviously generated data may be data generated from one or moreprevious applications of the present invention to the same patient forthe same tissue at prior times. This approach may be used to develop ahistory, to show the progression of the development of the clot forexample. Of course, the in vitro apparatus described herein could beused to carry out the same tests outside of the body, such as in alaboratory or a patient's home test kit.

Still further evaluation of the effectiveness of an anti-clottingtreatment may be performed, such as by evaluating the blood prior toapplication of the treatment by generating a series of acoustic pulsesand directing the series of pulses into the blood such that at least oneof the pulses is of sufficiently high intensity to induce physicaldisplacement of the blood, receiving at least two pulses reflected fromthe blood to establish a baseline and to estimate at least onecharacteristic of the physical displacement induced by the waves, andthen repeating these steps at least one time after administration of thetreatment Of course, as noted earlier, alternative sensing or receivingsteps may be taken to track the movement of the blood, such as by usingany of the alternative sensing techniques described above, e.g., laserDoppler, optical coherence tomography, etc. Repealed applications of thesteps at predetermined time intervals may be performed if needed toensure a stabilization of the properties measured, as a result of thetreatment. Alternatively, the analysis may indicate that a larger orsmaller dose of treatment is needed, or that the treatment isineffective for a particular patient.

Alternatively, evaluation of the effectiveness of an anti-clottingtreatment may be performed by carrying out the analysis steps a numberof times after treatment, at predetermined time periods after theadministration of the treatment, for example. The results generated fromeach iteration can then be compared and analyzed to note any changes inthe at least one physical characteristic that is beingmeasured/estimated.

Maintenance monitoring can be carried out by the same techniques noted,wherein a patient can be periodically tested to ensure that a clot hasnot progressed further and/or is dissolving.

FIG. 2 shows a schematic representation of an example of a system 50 forcharacterization of changes in physical properties of soft tissue overtime. In this example, a transducer 52, such as may be contained in adevice 10 as described above, or directly mounted, fixed to or integralwith a container holding a sample 51, for example, is connected to atransmitter 54 as well as receiver 56, both of which are controlled byprocessor 58 and timed by clock 60.

Clock 60 is provided to control the timing of application of radiationto the sample as generated by transmitter and converted to the acousticenergy at transducer 52, as well as the timing of receiving andinterpreting the reflected waves (echoes), by conversion throughtransducer 52 and receipt of the converted signals at receiver 56, allof which is controlled by one or more processors/microprocessors 58.

Displacements of the soft tissue may be induced by delivering one ormore acoustic pulses according to a predetermined frequency throughdevice 10. The displacements may be estimated by applying one or moresignal processing algorithms (e.g., minimum sum squared differencemotion tracking algorithm, etc.) to the acquired echoes of every nthdelivered pulse where “n” is a predefined integer. Alternatively, thesignal processing algorithms may be applied to every pulse received.Similarly, algorithms may be applied at every n^(th) time interval foroptical waves received. Parameter measurement may be initiated at apredetermined time after one or more coagulation reagents are added tothe sample, and such measurements may be repeatedly performed, e.g.,once after each passage of a pre-designated time period or according topre-defined time intervals for measurement. At each acquired time lapse,a time-displacement curve may be generated from which the viscoelasticparameters of the sample can be determined.

FIG. 3 is a graph 100 showing a set of time-displacement curves 110,120, 130 obtained during coagulation of a blood sample using thetechniques described. Curves 110, 120 and 130 are superimposed onaccompanying model predictions, where the mechanical properties of theforming thrombus are modeled by a modified Voigt model 150 as shown inFIG. 4. Experimental results and theoretical predictions show excellentagreement. The basis of the model from which the mechanical parametersare derived is the Voigt model in series with an inertial component.

The modified version 150 of the Voigt model may be used to model theviscoelastic response of blood to acoustic radiation force from whichmechanical parameters of the blood may be estimated. Model 150 includesan inertial component “m” in series with the traditional Voigt model,which includes a spring k in parallel with a dashpot μ, as shown in FIG.4. The governing differential equation for the model is:

$\begin{matrix}{{F(t)} = {{{kx}(t)} + {\mu \frac{}{t}{x(t)}} + {m\frac{^{2}}{t^{2}}{x(t)}}}} & (4)\end{matrix}$

where F(t) is the applied force as a function of time, x(t) is theinduced displacement as a function of time, k is the elastic constant, μis the viscous constant, and m is the inertial component. System 50applies radiation force by transmitting a series of pulses to the samelocation in the blood sample. Assuming that the pulse-to-pulse intervalis much shorter than the time constant of the blood's mechanicalresponse, the forcing function may be modeled as a temporal stepfunction as follows:

F(t)=Au(t)  (5)

where A is the force amplitude. Substituting equation (5) into equation(4) and solving for the displacement yields:

$\begin{matrix}{{x(t)} = {{\frac{\zeta = \sqrt{\zeta^{2} - 1}}{2\sqrt{\zeta^{2} - 1}}s{\square ^{{({{- \zeta} + \sqrt{\zeta^{2} - 1}})}\omega \; t}}} + {\frac{\zeta - \sqrt{\zeta^{2} - 1}}{2\sqrt{\zeta^{2} - 1}}s{\square ^{{({{- \zeta} - \sqrt{\zeta^{2} - 1}})}\omega \; t}}} + s}} & (6)\end{matrix}$

where ζ is the damping ratio, ω is the natural frequency (in radians persecond) and s is the static sensitivity. These parameters are definedas:

$\begin{matrix}{\zeta = \frac{\mu}{2\sqrt{k{\square m}}}} & (7) \\{\omega = \sqrt{\frac{k}{m}}} & (8) \\{s = \frac{A}{k}} & (9)\end{matrix}$

In the examples described herein, the force scaling constant A was notmeasured. Thus the time-displacement data in this situation can only beused to solve for relative parameters. To address this limitation, theequations (7), (8) and (9) are redefined according to the followingequations (10), (11) and (12) using relative measures of elasticityk_(r), viscosity μ_(r), and mass m_(r):

$\begin{matrix}{\zeta = \frac{\mu_{r}}{2\sqrt{k_{r}{\square m_{r}}}}} & (10) \\{\omega = \sqrt{\frac{k_{r}}{m_{r}}}} & (11) \\{s = \frac{1}{k_{r}}} & (12)\end{matrix}$

where k_(r)=k/A, μ_(r)=μ/A and m_(r)=m/A.

Although the viscosity, elasticity and inertia are measured asforce-dependent parameters, the natural frequency and the damping ratiostill remain force-free or force-independent parameters. It is furtherpossible to define a third force-independent parameter, i.e., the timeconstant τ as:

$\begin{matrix}{\tau = \frac{\mu_{r}}{k_{r}}} & (13)\end{matrix}$

The fact that the actual data shown in FIG. 3 waivers or oscillatessomewhat about the model data curves suggest that a different modelmight be used to even more closely model the behavior. In one possiblemodification, a dashpot would be placed in series with the model shownin FIG. 4. However, the model of FIG. 4 accurately described theresponse of the blood during formation of a clot with correlationbetween the data and the model of FIG. 3 being greater that 99% in mostof the cases analyzed.

EXAMPLE

The following example is put forth so as to provide those of ordinaryskill in the art with a complete disclosure and description of how tomake and use the present invention, and are not intended to limit thescope of what the inventors regard as their invention nor are theyintended to represent that the experiments below are all or the onlyexperiments performed. Efforts have been made to ensure accuracy withrespect to numbers used (e.g. amounts, temperature, etc.) but someexperimental errors and deviations should be accounted for. Unlessindicated otherwise, parts are parts by weight, molecular weight isweight average molecular weight, temperature is in degrees Centigrade,and pressure is at or near atmospheric.

An experimental system 1 as schematically represented in FIG. 5 wasused. Device 10 was mounted at one end of a rail and a container 30 wasmounted on the opposite end portion of rail 20. Device 10 included a 1.0cm diameter single piston transducer (General Electric Panametrics V327,Waltham, Mass.) mounted on a five-axis gimbal mount (NewportCorporation, Irvine, Calif.). The transducer had a fixed focus at 4 cm.The transducer was held with the focus at the center of a modified 4.5mL polystyrene cuvette 30 (Fisher Scientific) that held the bloodsample. Cuvette 30 was secured to the rail 20 at a slight tilt so thatreflections from the cuvette surface would be directed away from thetransducer. Each cuvette 30 was modified by drilling a hole 32 a,32 dapproximately 7 mm in diameter through the front and back sides ofcuvette 30 and using silicone sealant (Dow Corning, Baltimore, Md.) tomount a KAPTON® (Dupont, Wilmington, Del.) window over each opening. TheKAPTON® windows are acoustically transparent and were provided along theacoustic beam axis of the assembly. The assembly 1 was placed in a waterbath held at a room temperature of 21° C.

Transmitted pulses were Gaussian enveloped sinusoids with a centerfrequency of 10 MHz and a full-width half maximum fractional bandwidthof 75%. The sinusoidal pulses were amplified by 50 dB prior totransmission for a peak-to-peak amplitude of 136 volts. Based onhydrophone measurements performed in the lab, this transmit voltagecorresponded to an acoustic intensity (I_(spta)) of 300 mW/cm². A seriesof 4,000 acoustic pulses were transmitted by the transducer at a pulserepetition frequency of 5 kHz to generate acoustic radiation forcewithin the blood. The returning echoes of every tenth transmitted pulsewere acquired in order to estimate displacements induced by radiationforce. The same digital clock was used to drive pulse generation anddata acquisition, reducing sampling jitter to the order of picoseconds.

To confirm the accuracy of these techniques, preliminary experimentswere first performed with a control solution. Results obtained fromthese preliminary experiments were compared to results obtained from useof a conventional rheometer (TA Instruments AR-2000 constant stressrheometer, available from TA Instruments, Wilmington, Del.). An aluminumdouble-concentric cylindrical geometry was chose for the conventionalrheometer because it is best suited for lower viscosity samples that arenot sufficiently solid to maintain their structures under a cone andplate or parallel plate setup.

The control solution consisted of a liquid soap (Clean & Clear® DailyPore Cleanser, Johnson & Johnson) diluted with deionized water. Althougha broad variety of solutions were analyzed, including blood mimickingfluids (for flow measurements) and glycerol solutions, the liquid soapsolution specified above was bound to offer an appropriate viscoelasticresponse while remaining homogeneous and stable over time. Further, thespherical “micro-scrubbers” in the soap were excellent ultrasonicscatterers. The control solution consisted of 60% liquid soap dilutedwith 40% deionized water. A volume of 20 mL of the control solution wasprepared, subsequently vortexed, and placed overnight in a vacuumchamber at 20 mmHg to remove air bubbles trapped within the solution.

Approximately 4 mL of the control solution was placed in a 4.5 mLpolystyrene cuvette 30 (having been modified as described above) andsecured to rail 20 as described above. A sequence of 4,000 pulses wastransmitted according to the experimental protocol previously described.A sequence of ten acquisitions was obtained. The solution was gentlystirred with a needle between each acquisition to generate a new specklepattern and avoid settling of the “micro-scrubbers”. The remaining 16 mLof solution was used to perform a creep test in the conventionalcylindrical rheometer. Displacement data was obtained from an appliedshear stress. Because the amount of force applied by acoustic radiationcannot easily be quantified, the maximum displacement observed accordingto the present techniques (sonorheometry) in the control solution wasused to establish the applied shear stress for the creep test on theconventional rheometer. The computer controlling the conventionalrheometer was programmed to perform a sequence of ten repeated creeptests for each acquisition obtained with sonorheometry. The temperaturecontrol of the conventional rheometer was set to 21° C. to match thetemperature of the water bath during sonorheometry experiments.

Following the completion of the control experiments, sonorheometryexperiments were performed on blood samples of approximately 4 mL involume each. Blood was drawn from four healthy volunteers in theirmid-twenties and each sample was placed into a modified 4.5 mLpolystyrene cuvette 30 having been modified as described above. One ofthe four test subjects (referred to as “male 2”) indicated a recenthistory of a clotting disorder (deep vein thrombosis). Thrombin (0.5units per mL of blood) was immediately added to each drawn blood samplein order to induce coagulation. Each combined sample was invertedmultiple times to mix the thrombin within the sample. Each cuvette 30 ofcoagulating blood was mounted to the rail 20 as described above.Sonorheometry experimental protocol was initiated within two minutes ofthe addition of thrombin. Sonorheometry measurements were repeated overa seventy minute period in order to characterize the blood sample foreach respective cuvette 30. Data was acquired every minute for the firstten minutes, every two minutes for the next ten minutes, every threeminutes for the next fifteen minutes, and finally, every five minutesfor the next thirty-five minutes for a total of twenty-six acquisitionsper sample.

The sum squared differences (SSD) algorithm was applied between thefirst and the n^(th) echoes (where n is a predefined integer thatdefines how often echoes or waves are considered to estimate a tissuedisplacement measurement; in this example, n was set to 10) to determinethe tissue displacement at a given range, see Viola et al., “AComparison of the Performance of Time Delay Estimators in MedicalUltrasound”, IEEE Transactions on Ultrasonics, Ferroelectrics, andFrequency Control, vol. 50, no. 4, pp. 392-401, 2003, which isincorporated herein, in its entirety, by reference thereto. Sub-sampledelays were estimated by identifying the location of the peak of aparabolic fit about the minimum of the SSD grid. The ensemble of thesedisplacement estimates form a time-displacement curve that holdscombined information about both elastic and viscous components of theblood sample being analyzed.

In order to model the viscoelastic response of coagulating blood to thusestimate the mechanical parameters, the modified version of the Vogtmodel (described above with regard to FIG. 4) was applied to theexperimental data acquired. The governing differential equation for thisapplication is equation (1) above. Device 10 of system 1 appliedradiation force by transmitting a series of 4,000 ultrasonic pulses tothe same location within the blood sample.

A portion of the results obtained from analyzing the control solution isshown in FIGS. 6A-6C. Of one hundred paired time-displacement curves,each overlaying a curve obtained via sonorheometry over a correspondingcurve obtained with the convention cylindrical rheometer, three suchcurves are shown, see FIGS. 6A-6C, respectively. The sonorheometry datais presented as a continuous line 500 s, while the conventionalcylindrical rheometer data is displayed by circles 500 c. Similarresults to those shown were obtained for the remainder of the controlexperiments. The correlation coefficient between the data obtained bythe two methods ranged from 0.9990 to 0.9999, with an overall meancorrelation coefficient of 0.9993.

FIG. 7 shows estimated time constants (i.e., “tau” or “τ”) as a functionof maximum achieved displacement. Black circles 600 s represent the timeconstants estimated from the present methods (sonorheometry) while graydiamonds 600 c represent the estimates from the conventional rheometer.The time constants, as shown, have values ranging from approximately0.45 to 1.91 seconds, while the majority of the time constants liearound 0.6 seconds.

FIG. 8 shows a set of time-displacement curves obtained from one bloodsample along with the accompanying best fit model predictions. Thepredictions are the smooth lines and the time displacement curvesgenerated from the experimental data are the slightly wavering lines.The displayed curves indicate displacement at the same axial locationwith cuvette 30 for a single sample taken from female subject 2.Although coagulation data was acquired at twenty-six time intervals inthe seventy minute experimental period, for sake of simplicity, only theexperimental curves obtained at two, eight and twenty minutes have beendisplayed. Similar curves were obtained for the other acquisition times.Experimental results and best fit models show excellent agreement. Thepeak-to-peak error in displacement estimates was on the order of 0.5microns.

Maximum detected displacement of the blood samples progressivelydecreased as a blood clot started to form in each respective cuvette 30.After a certain amount of time, which was unique to each subject, noappreciable displacement could be detected. The results also suggestedthat the axial position from which displacements were detected becameprogressively narrower with the passage of time.

The estimated (calculated) relative modulus of elasticity and relativeviscosity values clearly showed that the relative modulus of elasticityincreased as the clot formed, and this was consistent for all foursubjects tested. As to viscosity, the two female subjects showedincreasing relative viscosity as the clots were forming, while for themale subjects, relative viscosity remained fairly constant, with someincrease near the conclusion of coagulation.

As to force-free (force independent parameters), the time constant,expressed in seconds, decreased with time for all four subjects, whichwas expected since the blood becomes stiffer as time elapses. Forexample, the time constant of the “female 1” subject decreased fromabout 0.6 seconds at the two minute test time to a value of about 0.3seconds at the twenty minute test time, and the time constant of the“male 2” subject had a value of 0.45 seconds at the two minute test timeand decreased to 0.2 seconds at the sixteen minute test time. Similartrends were observed for damping ratios, where clotting blood exhibitslower values. As expected, blood samples are always over-damped systems.In contrast with the other two force-free parameters examined, naturalfrequency increases as blood is coagulating, reaching values of about500 rad/sec (for female 1 and female 2).

1. A method of characterizing changes in at least one physical propertyof a biological material over time, said method comprising: generating aseries of acoustic pulses and directing said series of pulses into thebiological material such that at least one of the pulses is ofsufficiently high intensity to induce physical displacement of thebiological material; estimating at least one physical property of thebiological material based on measuring a displacement resulting fromsaid induced physical displacement of the biological material; andrepeating said generating and estimating steps after passage of a timeinterval.
 2. The method of claim 1, wherein said estimating is based onreceiving at least two of said acoustic pulses reflected from thebiological material and estimating the at least one physical propertybased on the acoustic pulses received.
 3. The method of claim 1, whereinsaid estimating is based on receiving optical reflections from thebiological material as the biological material is being physicallydisplaced and estimating the at least one physical property based on theoptical reflections received.
 4. The method of claim 1, wherein saidgenerating and estimating steps are repeated after passage of each of aplurality of predetermined time intervals.
 5. The method of claim 1,further comprising outputting at least one measured characteristic fromeach performance of said generating and measuring steps.
 6. The methodof claim 5, wherein said outputting comprises plotting time-displacementdata.
 7. The method of claim 1, wherein the biological material isblood.
 8. The method of claim 1, wherein said at least one physicalproperty comprises at least one parameter determined by fittingexperimental data including a plurality of said estimates ormeasurements, to a theoretical model defining the at least onecharacteristic.
 9. The method of claim 8, wherein said at least onephysical property comprises at least one of relative viscosity andrelative elasticity of the biological material.
 10. The method of claim1, wherein said at least one physical property comprises at least one ofrelative viscosity and relative elasticity of the biological material.11. The method of claim 1, wherein said at least one physical propertycomprises at least one force-free parameter.
 12. The method of claim 1,wherein said acoustic pulses are ultrasound pulses.
 13. The method ofclaim 7, further comprising calculating a characteristic of coagulationof the blood, based upon at least a portion of a time series of said atleast one physical property resultant from said generating and measuringsteps.
 14. The method of claim 7, further comprising identifying a stageof development of a blood clot in the blood, based upon at least aportion of a time series of said at least one characteristic resultantfrom said generating and measuring steps.
 15. The method of claim 1performed on a patient.
 16. The method of claim 15, wherein thebiological material is blood.
 17. The method of claim 1, furthercomprising receiving at least a portion of said pulses that pass throughthe biological material and estimating at least one acoustic property ofthe biological material.
 18. The method of claim 17, wherein said atleast one acoustic property comprises at least one of attenuation andspeed of sound.
 19. The method of claim 17, further comprisingestimating a magnitude of applied force of the at least one pulse havingsufficiently high intensity to induce physical displacement of thebiological material, based upon said at least one estimated acousticproperty.
 20. An apparatus for identifying changes in at least onephysical parameter of a biological material over time, said apparatuscomprising: an acoustic wave generator capable of repeatedly generatingacoustic pulses at least one of which is of sufficient intensity toinduce measurable physical displacement in the biological material; asensor adapted to sense at least one of optical waves and said acousticpulses after reflection by the biological material; a clock governingcycles during which said acoustic pulses are generated and during whichsensing of at least one of said acoustic and optical waves is carriedout; and a processor that receives input from said sensor and clock andcalculates data characterizing at least one characteristic of thephysical displacement induced over time.
 21. The apparatus of claim 20,further comprising at least one additional sensor adapted to be locatedon an opposite side of the biological material relative to a side of thebiological material from which said acoustic pulses are generated,wherein receipt of at least a portion of said waves that pass through atleast a portion of the biological material by said at least oneadditional sensor are measured to estimate at least one acousticproperty of the biological material.
 22. The apparatus of claim 20,wherein said acoustic wave generator comprises an ultrasonic transducer.23. The apparatus of claim 20, wherein an ultrasonic transducerfunctions both as said acoustic wave generator and as said sensor. 24.The apparatus of claim 20, wherein said sensor comprises at least oneoptical sensor.
 25. The apparatus of claim 20, further comprising meansfor outputting said data.
 26. The apparatus of claim 25, wherein saidmeans for outputting outputs time-displacement data.
 27. The apparatusof claim 20, wherein the biological material is blood and wherein saidat least one characteristic comprises at least one of relative viscosityand relative elasticity of the blood.
 28. The apparatus of claim 20,wherein the biological material is blood, said system further comprisingmeans for calculating a rate of coagulation of the blood, based upon atleast a portion of a series of said data.
 29. The apparatus of claim 20adapted to non-invasively deliver said acoustic pulses into a patientfrom a location outside of the patient.
 30. The apparatus of claim 20adapted such that at least a portion of said apparatus is deliverableinvasively within a patient.
 31. The apparatus of claim 30, wherein thedelivery is intravascular.
 32. The apparatus of claim 30, wherein thedelivery is intracavity.
 33. The apparatus of claim 20, wherein saidacoustic wave generator comprises at least one component selected fromthe group consisting of: piezoelectric transducers, electric circuits togenerate rapid heating, an ultrasonic generator fabricated usingmicroelectromechanical systems (MEMS), capacitive micromachinedultrasound transducers, PVDF transducers, and a laser to heat a targetmaterial.
 34. The apparatus of claim 20 applied directly in contact withthe biological material.
 35. The apparatus of claim 34, wherein thebiological material is blood.
 36. A system comprising the apparatus ofclaim 20, and at least one sample container for containing a sample foranalysis by said apparatus.
 37. The system of claim 36 comprising aplurality of said sample containers for parallel processing of multiplesamples in said plurality of sample containers. 38-42. (canceled)
 43. Amethod of characterizing a stage of development of a blood clotcomprising carrying out the method of claim 36, and comparing the atleast one estimated characteristic to previously generated data to gaugethe stage of development of the blood clot. 44-46. (canceled)
 47. Themethod of claim 1, wherein the biological material comprises softtissue.
 48. The method of claim 1, wherein the biological materialcomprises blood.
 49. The method of claim 1, comprising placing thebiological material in contact with a control material, and controllingthe temperature of the control material.
 50. The apparatus of claim 20,wherein said means for controlling temperature of the biologicalmaterial being physically displaced comprises a temperature-controlledsecond material, said temperature-controlled second material beingplaceable in contact with the biological material being physicallydisplaced to control the temperature of the biological material beingphysically displaced.
 51. The method of claim 1 performed non-invasivelyon a patient.
 52. The apparatus of claim 20, wherein the biologicalmaterial comprises soft tissue.
 53. The apparatus of claim 20, whereinthe biological material comprises blood.
 54. A method of characterizingchanges in at least one physical property of a material over time, saidmethod comprising: applying a device to an animal body comprising thematerial; generating a series of acoustic pulses and directing saidseries of pulses into the material such that at least one of the pulsesis of sufficiently high intensity to induce physical displacement of thematerial; estimating at least one physical property of the materialbased on measuring a displacement resulting from said induced physicaldisplacement of the material; and repeating said generating andestimating steps after passage of a time interval.
 55. The method ofclaim 54, wherein said applying a device comprises inserting at least aportion of the device invasively within the body.
 56. The method ofclaim 54, wherein the material comprises soft tissue.
 57. The method ofclaim 54, wherein the material comprises blood.
 58. An apparatus foridentifying changes in at least one physical parameter of a biologicalmaterial over time, said apparatus comprising: an acoustic wavegenerator capable of repeatedly generating acoustic pulses at least oneof which is of sufficient intensity to induce measurable physicaldisplacement of the material; a sensor adapted to sense at least one ofoptical waves and said acoustic pulses after reflection by thebiological material; a temperature controller configured to controltemperature of the biological material being physically displaced; aclock governing cycles during which said acoustic pulses are generatedand during which sensing of at least one of said acoustic and opticalwaves is carried out; and a processor that receives input from saidsensor and clock and calculates data characterizing at least onecharacteristic of the physical displacement induced over time.
 59. Theapparatus of claim 58, wherein the biological material comprises softtissue.
 60. The apparatus of claim 58, wherein the biological materialcomprises blood.
 61. An apparatus for characterizing at least onephysical property of a biological material, said apparatus comprising:an acoustic wave generator configured to generate a series of acousticpulses and direct said series of pulses into the biological materialsuch that at least one of the pulses is of sufficiently high intensityto induce physical displacement of the material, and wherein each ofsaid acoustic pulses is of finite duration and of brief durationrelative to a duration of said generating and said directing; atemperature controller operable to control a temperature of thebiological material; a sensor configured to receive pulses reflected bythe biological material; and a processor configured to receive signalsfrom said sensor and measure a displacement, either directly orindirectly, of the material resulting from said induced physicaldisplacement thereof, wherein the temperature of the material iscontrolled over a duration of said measuring a displacement, andestimate at least one characteristic of the physical displacement basedon said measuring.
 62. A method of characterizing at least one propertyof a biological material, said method comprising: testing a firstportion of the biological material; estimating a quantitative value of aproperty of the first portion; treating a second portion of thebiological material with a treatment to vary a quantitative value ofsaid property from said quantitative value of the property of the firstportion having been estimated; testing the second portion of thebiological material having been treated; estimating a secondquantitative value of said property of the second portion; comparingsaid first quantitative value with said second quantitative value; andevaluating an effect of said treating on said property of the biologicalmaterial.
 63. The method of claim 62, wherein said testing a firstportion of the biological material and said testing the second portionof the biological material are performed in parallel.
 64. The method ofclaim 62, further comprising: treating a third portion of the biologicalmaterial with a treatment to vary a quantitative value of said propertyfrom said first and second quantitative values of the property of thefirst and second portions having been estimated; testing the thirdportion of the biological material having been treated; estimating athird quantitative value of said property of the third portion;comparing said third quantitative value with said first and secondquantitative values; and evaluating an effect of said treating on saidproperty of the biological material.
 65. The method of claim 62, whereinthe biological material is blood.
 66. The method of claim 65, whereinsaid treating comprises treating the second portion of the blood with ananti-clotting treatment.
 67. The method of claim 65, wherein saidtreating comprises treating the second portion of the blood with apro-clotting treatment.
 68. The method of claim 62, wherein said testingcomprises: generating a series of acoustic pulses and directing saidseries of pulses into the biological material such that at least one ofthe pulses is of sufficiently high intensity to induce physicaldisplacement of the biological material; and measuring a displacement,either directly or indirectly, of the biological material resulting fromsaid induced physical displacement thereof; and wherein said estimatingis based on said measuring.
 69. The method of claim 68, wherein thetemperature of the biological material is controlled over a duration ofsaid measuring a displacement.
 70. A method of characterizing a stage ofdevelopment of a blood clot, said method comprising: generating a seriesof acoustic pulses and directing said series of pulses into blood inwhich the blood clot is to be characterized, such that at least one ofthe pulses is of sufficiently high intensity to induce physicaldisplacement of the blood, and wherein each of said acoustic pulses isof finite duration and of brief duration relative to a duration of saidgenerating and said directing; controlling a temperature of the blood;measuring a displacement, either directly or indirectly, of the bloodresulting from said induced physical displacement thereof, wherein thetemperature of the blood is controlled over a duration of said measuringa displacement; and estimating at least one characteristic of thephysical displacement based on said measuring.
 70. The method of claim7, further comprising calculating a rate of fibrinolysis of the blood,based upon at least a portion of a time series of said at least onecharacteristic resultant from said generating and measuring steps. 71.The method of claim 7, further comprising calculating progress ofhemostasis of the blood, based upon at least a portion of a time seriesof said at least one characteristic resultant from said generating andmeasuring steps.
 72. The method of claim 1, wherein said at least onephysical property comprises viscosity of the biological material. 73.The method of claim 1, wherein said at least one physical propertycomprises elasticity of the biological material.
 74. The method of claim1, wherein said at least one physical property comprises a combinationof shear and compressive modulus of the biological material.
 75. Themethod of claim 13, wherein said characteristic comprises a rate. 76.The method of claim 13, wherein said characteristic comprises a clotstiffness.
 77. The method of claim 13, wherein said characteristiccomprises a time to clot.
 78. The method of claim 13, wherein saidcharacteristic comprises a time to clot dissolution.
 79. The method ofclaim 17, wherein said pulses that pass through the biological materialare received on an opposite side of the biological material from a sidewhere said pulses enter the biological material.
 80. The system of claim37, wherein different ones of said plurality of sample containerscontain different reagents, relative to one another for inhibiting orexciting at least one coagulation characteristic.